Controlled transfer biological sample collection devices and methods of using such devices

ABSTRACT

The field of the present invention pertains to a controlled transfer biological collection device using a dry solid storage and transfer medium and a method for the collection of biological material of interest (genetic or proteinaceous material) in a form suitable for storage and/or subsequent analysis. Specifically, the present invention provides for a sampling device that controls the transfer of the biological sample to the storage medium by holding the storage medium and a moveable sample collection member having an analyte collection surface. The invention further provides for a method not only for storing a biological analyte on this collection device but also for analyzing the stored biological analyte using methods that are suited for automated analyzing systems.

CROSS-REFERENCES TO RELATED APPLICATIONS

The present application is a continuation-in-part application of U.S.Ser. No. 11/707,313, filed Feb. 16, 2007, the disclosure of which isincorporated herein by reference.

BACKGROUND OF THE INVENTION

(1) Field of the Invention

The field of the present invention pertains to a controlled transferbiological collection device using a dry solid storage and transfermedium and a method for the collection of biological material ofinterest (genetic or proteinaceous material) in a form suitable forstorage and/or subsequent analysis. Specifically, the present inventionprovides for a sampling device that controls the transfer of thebiological sample to the storage medium by holding the storage mediumand a moveable sample collection member having an analyte collectionsurface. The invention further provides for a method not only forstoring a biological analyte on this collection device but also foranalyzing the stored biological analyte using methods that are suitedfor automated analyzing systems.

(2) Description of the Related Art, Including Information DisclosedUnder 37 CFR 1.97 & 1.98

The collection of biological samples (such as blood) and extracting DNAfor genetic analysis from the sample has been widely used by theforensics and medical community for identification purposes, forpaternity testing , for genetic diagnostic testing in new born screeningprograms, for genetic typing for predisposition to disease and forgenetic characterization for drug susceptibility. However, due to theinvasive nature of blood collection, alternative non-invasive methodsare coming into favor. Current methods involve scraping cellular mucosafrom inside the oral cavity using any of a number of different devicessuch as cytobrushes, cotton or Dacron swabs, mouthwash swish and rinsemethods, foam tipped swabs, and supported cellulosic filter papercollection techniques (known as the Bode method). These methods requiretime-consuming, labor intensive extraction methods.

The introduction of treated storage matrices into the forensicscommunity has significantly streamlined the collection and extraction ofDNA from a variety of samples. The use of FTA® brand treated matrices(from Whatman, Inc. of Florham Park, N.J. USA) with non-invasive buccalcell collection techniques presents a new set of problems. With the useof conventional buccal swabs, one can fail to transfer buccal cells tothe treated matrix in a consistent and reproducible manner. If the swabused to collect the sample is separate and distinct from the treatedmatrix receiving the sample, then forensic traceability issues arise,particularly if the two become separated later in the chain of custodyof forensic evidence.

Examples of treated matrices for biological sample collection or storageand associated collection devices can be found in the following USpatents: U.S. Pat. No. 6,627,226, U.S. Pat. No. 6,447,804, U.S. Pat. No.6,294,203, U.S. Pat. No. 6,168,922, U.S. Pat. No. 5,976,572, U.S. Pat.No. 5,972,386, U.S. Pat. No. 5,939,259, and U.S. Pat. No. 5,756,126.Basically, these patents use two different methodologies for stabilizingbiological samples.

The first stabilizing method uses a combination of an absorbent materialas a storage medium that does not bind to nucleic acids and a chaotropicsalt impregnated about the storage medium. (For the purposes of thecited prior art and the present invention, “chaotropic salts” includeany substance capable of altering the secondary, tertiary, or quaternarystructure of biomolecules in aqueous solution, but leaves the primarystructure intact.) Preferably, a chaotropic salt is said to inactivateany nucleic acid amplification inhibitors present in the biologicalsource, by precipitation, by inducing the inhibitor to irreversibly bindto the matrix, or by causing substantially irreversible denaturation ofthe inhibitor. Suitable chaotropic salts include guanidinium salts suchas guanidine isothiocyanate, guanidine thiocyanate, guanidinehydrochloride, sodium iodide, sodium perchlorate, potassium iodide,sodium isothiocyanate, urea, or combinations thereof.

The second stabilizing method also uses a dry solid storage medium but adifferent adsorbed or absorbed stabilizer. Here, the protectantcomposition comprises a protein denaturing agent (such as an anionicdetergent) and a free radical trap (such as a weak base, and a chelatingagent, and optionally, uric acid or a urate salt).

BRIEF SUMMARY OF THE INVENTION

The present invention relates to a controlled transfer biologicalcollection device using a dry solid storage and transfer medium and amethod for the collection of biological material of interest (genetic orproteinaceous material) in a form suitable for storage and/or subsequentanalysis.

The present collection device for a biological sample that containsdegradable biologically sourced analytes comprises three elements. Amoveable sample collection member is one element and is equipped with ananalyte collection surface that, preferably, has the ability to absorbmore sample than is necessary for transfer to a storage medium. Astorage medium suitable for collecting and storing the biological sampleis held in place by a storage medium holder. The holder not only keepsthe operator's fingers away from the storage transfer location, but alsoprovides a holding means for holding the storage medium in a fixedposition and for applying contact pressure between the storage mediumand the analyte collection surface. The holder also has a means forholding the moveable sample collection member. Thus, the two elements,the storage medium and the analyte collection surface are held togetherfor traceability purposes.

Functionally, the moveable sample collection holding means allows themoveable collection member, and its analyte collection surface, to movebetween a first open position for collecting the biological sample onthe analyte collection surface prior to sample collection and a secondclosed position facing or contacting at least a portion of the storagemedium after collection and transfer of the sample. For the purposes ofthe present invention, the term “surface” refers to more than atwo-dimensional space, including volume as well. Thus, a “surface” canbe the volume of a foam pad, for example, and not just its contactsurface area.

In use, one takes the above described device and contacts the analytecollection surface with the biological sample. The moveable collectionmember is moved towards the storage medium such that the analytecollection surface and the storage medium are brought in contact,allowing the transfer of the biological sample to the storage medium.

In preferred embodiments, one engages the holding means on the storagemedium holder in doing so, thereby allowing the analyte collectionsurface to be held facing the storage medium after the transfer iscomplete.

Preferably, the means for holding the moveable sample collection membercomprises a resilient member, which may be molded into the storagemedium holder. In one embodiment, the resilient member is such that, inuse, it engages the moveable collection member and releasably holds itin a fixed position such that the surface of the moveable collectionmember is in contact with the storage medium, thereby maintaining auniform and constant pressure between the moveable collection membersurface and the storage medium. Preferably the pressure is sufficient tofacilitate the transfer of the biological sample from the analytecollection surface of the moveable sample collection member to thestorage medium. The person skilled in the art will be able to determinethe relative position of the resilient member which is required in orderto ensure sufficient transfer of the biological sample from the analytecollection surface of the moveable sample collection member to thestorage medium.

For analysis of the biological sample, the storage medium is manipulatedso as to remove at least a portion of the biologically sourced analytepresent on the storage medium.

Examples of storage media suitable for the present invention includeuntreated filter paper, such as #903® brand paper (Whatman, Inc.,Florham Park, N.J. USA) or treated filter papers, such as FTA and FTAElute brand paper (also from Whatman, Inc., Florham Park, N.J. USA).These treated matrices are described in US patents referenced above.Such treated matrices provide a simple safe method for collection,shipping and storage of biological samples. They also containchemistries which make it easy to isolate DNA from complex samples suchas blood. Samples collected on treated or untreated matrices are driedfor storage and can be stored at room temperature for long periods oftime.

An object of the invention is to provide a controlled transfer of abiological sample to a dry, treated solid storage and transfer medium,such as providing a reproducible pressure or movement between theanalyte collection surface and the storage medium.

A second object of the invention is to provide a device or method thathas a spare sample source in sample retained in an absorbent analytecollection surface.

A third object of the invention is to provide a device or method thatretains the sample collector surface and the storage medium together forchain of custody traceability purposes.

A fourth object of the invention is to provide a device or method inwhich the storage medium can be processed by automated analyzingmethods.

A fifth object of the invention is to provide a device or method for thelong term storage for biological samples.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a perspective view of a preferred embodiment of the presentinvention showing the claimed element in a closed position;

FIG. 2 is a perspective view of a preferred embodiment of the presentinvention showing the claimed element in an open position;

FIG. 3 is a plan and sectional view of the FIG. 1 device;

FIG. 4 is a plan and sectional view of the FIG. 2 device;

FIG. 5 is a perspective view of a further embodiment of the presentinvention showing the claimed element in an open position;

FIG. 6 is a perspective view of a further embodiment of the presentinvention showing the claimed element in a closed position;

FIG. 7 is a graph illustrating the PCR product concentrations obtainedfrom β-globin PCR amplifications following use of THP-1 cells in adevice of the present invention;

FIG. 8 a is a punch map showing the buccal cell application area aftertransfer of buccal cells using a device according to the presentinvention;

FIG. 8 b illustrates the distribution of buccal cells on the storagemedium using the method of the present invention;

FIG. 8 c is a box and whisker plot of PCR product concentrationsobtained after β-globin PCR amplifications from 11 discs punched fromeach FTA storage medium;

FIG. 9 a is a comparison of the transfer patterns obtained using themethod of the present invention and the traditional swab method;

FIG. 9 b is a comparison between the method of the present invention andswab methods for the collection of buccal cells from five differentdonors; and

FIG. 10 shows the results of an STR analysis of four donor individualsusing Promega PowerPlex 16.

DETAILED DESCRIPTION OF THE INVENTION

A preferred embodiment is shown in FIG. 1. The collection device (10)for a biological sample that contains degradable biologically sourcedanalytes comprises a moveable sample collection member (20) having ananalyte collection surface (22), a storage medium (30), and a storagemedium holder (40) having a means for holding the storage medium in afixed position (50) and a means for holding the moveable samplecollection member (60). The moveable sample collection holding meansallows the moveable collection member surface to move either from afirst closed position facing or contacting at least a portion of thestorage medium (as shown in FIGS. 1 and 3) to a second open position forcollecting the biological sample on the analyte collection surface (asshown in FIGS. 2 and 4) or vice versa.

Preferably, the means for holding the moveable sample collection membercomprises a unitary connection between the storage medium holder and themoveable sample collection member (as shown in the FIGURES). Alsopreferably, the moveable sample collection surface is dimensioned andconfigured to be in spring tension away from the storage medium surfacewhen held by the member holding means such that the analyte collectionsurface is held off the storage medium thereby allowing enough space forair drying of the storage medium after transfer of the sample to thestorage medium from that surface.

Preferably the storage medium will also comprise at least onestabilizing reagent that preserves at least one biological sampleanalyte for transport or storage. Suitable such reagents include eitherthe combination of a weak base, a chelating agent, and, optionally, uricacid or a urate salt or simply the addition of a chaotropic salt, aloneor in combination with a surfactant.

The “weak base” of the composition may be a Lewis base which has a pH ofabout 6 to 10, preferably about pH 8 to 9.5. One function of the weakbase is to act as a buffer to maintain a composition pH of about 6 to10, preferably about pH 8.0 to 9.5, for example, pH 8.6. Hence, a weakbase suitable for the composition of the invention may, in conjunctionwith other components of the composition, provide a composition pH of 6to 10, preferably, about pH 8.0 to 9.5. Suitable weak bases according tothe invention include organic and inorganic bases. Suitable inorganicweak bases include, for example, an alkali metal carbonate, bicarbonate,phosphate or borate (e.g., sodium, lithium, or potassium carbonate).Suitable organic weak bases include, for example, tris-hydroxymethylamino methane (Tris), ethanolamine, triethanolamine and glycine andalkaline salts of organic acids (e.g., trisodium citrate). A preferredorganic weak base is a weak monovalent organic base, for example, Tris.The Tris may be either a free base or a salt, for example, a carbonatesalt.

A preferred chelating agent is a strong chelating agent. By “strong”chelating agent it is meant that the agent binds multivalent metal ionswith a comparable or better affinity than ethylene diamine tetraaceticacid (EDTA). A preferred chelating agent according to the invention isEDTA.

Anioinic surfactants are examples of surfactants which are useful in thepresent invention. A preferred anionic detergent is a strong anionicdetergent. As used herein, a “strong” anionic detergent includes ahydrocarbon moiety, aliphatic or aromatic, containing one or moreanionic groups. Particularly preferred anionic detergents suitable forthe invention include sodium dodecyl sulphate (SDS) and sodium laurylsarcosinate (SLS). In a preferred embodiment, the anionic detergentcauses inactivation of most microorganisms which have protein or lipidsin their outer membranes or capsids, for example, fungi, bacteria orviruses. This includes microorganisms which may be pathogenic to humansand are present in a biological sample.

Also preferably, the storage medium will have a visual delineation (32)placed around the transfer area of the storage medium such that ifremoved from the storage holding means an operator can know where thematerial was deposited without reference to the device.

The present device can be used to collect degradable biologicallysourced analytes such as nucleic acids, proteins, and respectivefragments thereof. The biological sample can be selected from the groupconsisting of saliva, blood, serum, lymph fluids, buccal cells, mucosalcells, cerebrospinal fluid, semen, vaginal fluid, feces, plasma, urine,a suspension of cells, or a suspension of cells and viruses.

Preferably, the present device is dimensioned and configured such thatthe storage medium holder (40) releaseably holds the storage medium (30)in the fixed position by the holding means (50) (such as the plasticarms shown in the FIGURES). Thus, one can separate the storage mediumfrom the storage holder for subsequent processing or storage. Thetension on the storage medium should allow for manual or automatedextraction, but not allow for accidental loss of the storage medium fromthe device.

As described above, the means for holding the moveable sample collectionmember may comprise a resilient member (70) positioned on the storagemedium holder. This is illustrated in FIG. 5. In use, where the moveablesample collection member is moved to a closed position, the resilientmember on the storage medium holder engages the moveable collectionmember and releasably holds it in a fixed position such that the surfacethereof is in contact with the storage medium, thereby maintaining auniform and constant pressure between the moveable collection membersurface and the storage medium.

In some cases, one can dimension and configure the storage medium holderso as to expose at least a portion of the storage medium for removal ofthe storage medium from the storage medium holder.

Preferably, the analyte collection surface (22) comprises an absorbentmaterial, such as a conventional porous polyurethane foam pad (fromPowell Products, Inc. of Colorado Springs, Colo. USA), that is suitablefor collecting a biological sample. Examples of other suitable absorbentmaterials include hydrophilic non-reticulated, closed cell foams,hydrophilic, non-reticulated, open cell foams, hydrophobicnon-reticulated closed cell foams, hydrophobic non-reticulated open cellfoams, hydrophobic reticulated open cell foams, absorbent gel materialsused for transfers and cotton based absorbent material. The analytecollection surface should be dimensioned and configured such that thevolume of sample is controlled. By controlling the volume, anystabilizing reagents on the storage medium are not overloaded in theirrespective protecting capacity. If used in buccal swab applications, thepad should be dimensioned and configured to fit within the human mouth.

For record keeping and traceability the present device should alsocomprise an identification label (such as conventional bar coding) onnot only the storage medium, but also the collection member, and if notunitary, the storage medium holder as well.

To ensure device integrity, the present device can also comprise asterility envelope surrounding the other device elements. Preferably,those other elements are sterile and free from any biological sampleanalytes (made for example, from medical grade plastics), which can bedone through conventional techniques such as irradiation after theenvelope is sealed.

Kits can be made that incorporate the above device along with anycombination of associated equipment or reagents including purificationreagents, buffers, or the like and storage systems, containers, or thelike.

In this regard, the present invention further provides a kit comprisinga device as defined herein and one or more components selected from thegroup consisting of purification reagents for subsequent analysis of thesample, buffers, storage systems and containers.

Example of Device Use

The present device can be used for biological sample collection for thefollowing purposes: the collection of buccal cell samples for criminaldatabases; the collection of crime scene samples (i.e., rehydratedblood, semen, saliva and liquid samples of the same); the collection ofsexual assault samples; the collection of buccal samples for populationgenetics or pharmacogenomics studies; the collection of buccal samplesfor personal genetic ID archiving; the collection of bacterial orparasite samples from food sources; the collection of blood from meat atslaughterhouse for meat traceability; or the collection of biologicalsamples from animals for veterinary diagnostics.

EXAMPLES Example 1 Cell Transfer Assays

Cell transfer assays were performed in order to evaluate the transferefficiency of the device of the present invention. The device used ineach of these experiments was a device as illustrated in FIG. 5. Theanalyte collection surface was formed from foam (Aquazone® availablefrom Reilly Foam Corporation, Pa., USA) and the storage medium comprisedFTA filter paper (obtained from Whatman, Inc., N.J., USA).

THP-1 cultures were grown to densities of 10⁶ cells/ml, centrifuged andsubsequently resuspended at a concentration of 10⁷ cells/ml. A serialdilution of this stock was performed to give concentrations of 10⁵, 10⁴and 10³ cells/ml. Each of these dilutions was applied to the foamcollection surface of a device according to the present invention (100μl each); the devices were then closed and clipped in place for 10seconds before release of the applicator foam to the resting position.

2 mm discs were punched from the white application area of the pinkindicator FTA filter paper card and placed into 0.5 ml tubes with 200 μlFTA Purification Reagent. Following a five minute incubation at roomtemperature, the tubes were finger-flicked for 10 seconds. Liquid wasremoved and the wash step repeated two more times for a total of threewashes with FTA Purification Reagent. TE buffer (200 μl ) was added toeach tube, incubated for 5 minutes at room temperature, and the TE wasremoved and discarded. This was repeated for a total of two washes usingTE buffer, after which the disc was dried for 1 hour at roomtemperature.

PCR amplification of the β-globin gene and capillary electrophoresiswere then performed. Analysis was performed using β-globin PCR assaysvia Exeprion Bioanalyzer 1K DNA chips. DNA detection was defined as thepresence of β-globin amplicons on the Experion 1K Chips with a detectionlimit at 0.1 ng/μl.

The results obtained are set out in FIG. 7 which illustrates thatdetectable amplification was observed using all concentrations of cells.More specifically, PCR product was successfully detected even followingapplication of only 100 cells to each device. Excellent results wereobtained where 1000 cells were applied to the applicator foam to yieldβ-globin PCR amplicon concentrations ranging from 1.5 to 3.2 ng/μl.

Example 2 Buccal Cell Mapping Experiments

Samples were collected from four subjects, each using two devices asillustrated in FIG. 5. The samples were collected according to thefollowing protocol:

a) Holding the plastic stem above the hinge joint, the foam tip of thedevice was placed in the mouth of the subject and the foam analytecollection surface was rubbed on the inside of the cheek for 15 seconds.This procedure was repeated using the opposite cheek. The foam analytesurface was rubbed along the gum-line, at the fold line of the cheek andunder the tongue, soaking up as much saliva as possible. The foamanalyte surface was then removed from the mouth.

b) The protective film of the storage medium was removed, exposing theFTA Card storage medium.

c) The device was folded at the hinge joint and the foam analyte surfacewas pressed onto the FTA Card making sure that the foam samplecollection surface was held in place by the clip at the front of the FTACard holder. The device was then closed and the sample collection memberwas pushed into the lowest position on the clip. The device was left inthis position for 10 seconds.

d) Whilst holding the FTA Card holder, the device handle was bent backto release the foam sample collection surface from the clip and to pullthe sample collection member up to the top position on the clip. Thedevice was bent to lift the foam sample collection surface from the FTACard storage medium.

e) The FTA card storage medium was removed and dried for 3 hours, readyfor subsequent analysis.

Eleven discs (2.0 mm diameter) were punched from each card as shown inFIG. 8 a, and PCR amplifications targeting a 268 by portion of theβ-globin gene were completed and are shown in FIG. 8 b. The ampliconswere quantified using an Experion analyzer (Bio-Rad) and are showngraphically in FIG. 8 c as a Box and Whiskers Plot. A total of 88punches were analyzed from eight Indicating FTA cards; 87 containedsufficient DNA template for successful PCR, i.e. only 1 failure in 88punches (in this case, the failed punch was taken from the periphery ofthe application area). Negative controls included paper-only samples(devoid of biological sample) and water only (no template PCR). In bothnegative control cases, amplification of PCR products was not detected.

With reference to FIG. 8 a, the punch maps show the buccal cellapplication area after transfer of buccal cells from the foam applicatorto the Indicating FTA card (storage medium) using the device of thepresent invention. Eleven 2 mm punches were removed from each card asshown in FIG. 8 a. The large circle represents the typical area of celltransfer indicated by a change in Indicating FTA color from pink towhite.

With reference to FIG. 8 b, the concentration of DNA detected isdirectly proportional to the number of buccal cells transferred to theFTA card. Four different donors used device of the present invention toharvest buccal cells from inside their cheeks and transfer them toIndicating FTA cards for analysis. Panels A-D show DNA distribution mapsof DNA on the FTA card after transfer of cells from the devices. Eachdata point represents the yield of PCR amplicons (ngs) produced from theβ-globin amplification of DNA present on a 2 mm disk. The PCRamplification was quantified using the Experion Bioanalyzer. Thedistribution of DNA present on the cards indicates that the transfer ofcells was consistently uniform.

With reference to FIG. 8 c, the range is shown as the whiskers and thebox shows the area where 50% of the data points lie. The bar in the boxrepresents the median value for the 11 data points. The data shown inthe Box and Whiskers Plot indicates that the transfer of buccal cellsfrom the sample collection surface to the storage medium was consistent.Sample 1B contains a single failed PCR reaction; the only one out of 88PCR amplifications. Sample 2A contained a single outlier point thatskewed the upper range of PCR yields, and this is likely the result of acell-clump transferred to the FTA card. PCR products showed minorvariation in concentration within an individual set, but the variationis most notable between subjects; for example subject #4 harvestedsubstantially more cells than other subjects on both collection devicesused. This serves to underscore the differences in shedding buccal cellsfrom one person to the next.

The final set of experiments utilizing buccal cell maps are displayed inFIGS. 9 a and 9 b, repsectively. These figures show the results of acomparative analysis of the method of the present invention against thetraditional swab method of buccal cell collection. Five buccal celldonors collected two buccal cell FTA cards using the method describedabove and two cards using the traditional swab method. The protocol forcell collection identified above was followed except that the swab wasrolled onto the FTA card (storage medium).

Each donor was sampled a total of four times (A-D), using two devices ofthe invention and two swabs. To remove sampling order bias, thecollection method order was varied between individuals in the ordershown in FIG. 9 b (A=1^(st) to D=4^(th)). Following sample collection,FTA cards were processed using standard protocols as described above andPCR was performed to amplify a fragment of the β-globin gene from 11positions within the application area as described above. Resultant PCRproducts were quantified using an Experion Bioanalyzer.

The comparison of performance between the device of the presentinvention and the traditional swab method reveals that the presentmethod provides for a well defined area of buccal cell transfer whilethe swab produces variable transfer patterns that require the user tomake a “best guess” of where to collect a punch sample (FIG. 9 a). Thedevice of the present invention provides a well defined application areamaking automated punching more accurate. The Box and Whiskers Plot inFIG. 9 b shows that the method using the device of the present inventionconsistently collects higher yields of buccal cells and generates a moreuniform transfer of cells than does the swab method regardless of theorder in which samples were collected.

Example 3 STR Analysis

Buccal samples utilized for STR analysis were taken from the samecollection devices in Example 2 above. Two 1.2 mm punches were collectedfrom central locations of each Indicating FTA card storage medium. Allpunches were washed and dried following the protocol outlined in Example1.

STR analysis was performed using the Promega PowerPlex 16® systemfollowing the manufacturer's instructions. A processed and driedIndicating FTA punch was used in each PCR reaction as a method of directamplification from the punch. PCR was carried out on an AppliedBiosystems 7900HT, and PCR products were visualized on an AppliedBiosystems 310 Genetic Analyzer. Analysis of products was carried outwith GeneMapper 3.2® software.

All 4 sets of buccal collection produced excellent quality results forall 16 alleles above 250 RFU's (relative fluorescent units) as seen inFIG. 10. This exceeds the desired criterion from the Design Inputdocument of 200 RFU's. Although, there was some minor variation in theintensity of the peaks, the peak balance and peak intensity remainedwell within the GeneMapper software acceptable parameters. As expected,replicate devices from the same individual produced the same allelicprofile. Allelic profiles are found in Table I. A series of 50 sampleswere collected with the device of the present invention, processed andanalyzed using Promega's PowerPlex 16 system of STR analysis (Table 2).

TABLE 1 D3S1358 TH01 D21S11 D18S51 Penta_E D5S818 D13S317 D7S820 Donor 117   6/9.3 28/29 13/15 12/14  8/11 12 10/11 Donor 2 14/16 7/8 30/31 17 7/12 10/11  8/12 10/11 Donor 3 16/17 9.3 30/31 12/16  5/13 11/12 1111/12 Donor 4 15/17   9/9.3 29/30 15/20  7/15 11/13 11/12  8/12 D16S539CSF1PO Penta_D AMEL vWA D8S1179 TPOX FGA Donor 1 11/13 10/12 10/13 X/Y18 13  9/11   22/22.3 Donor 2  9/13 12/13 10/13 X/Y 17/18 14/17  8/1120/24 Donor 3 11/14 11/12 9 X 15/17 14  8/11 21/23 Donor 4 11/13 10/12 9/13 X 16/17 11/14  8 19/21

TABLE 2 number of Promega's PowerPlex 16 STR System samples AmelogeninCODIS alleles Penta D & E Totals Total possible allele calls 50 50 650100 800 Total correct allele calls* 49 49 642 98 789 Total failed allelecalls 1 1 8 2 11 Re-run of failed sample correct allele 1 1 13 2 16calls** Total Correct Allele calls after sample re- 50 50 650 100 800run % accuracy after first set of injections 98.6% % accuracy afterfailed sample re-run  100% *all allele calls of peaks with ≧200 RFU **10second reinjection of sample; peak balance was <60% for severalheterozygous peaks

In FIG. 10, A, B, C, and D are results from buccal cell donors 1, 2, 3,and 4 respectively. The red line indicates the point of 250 rfu's. BothPenta D and E alleles are marked as indicators for DNA template qualitysince they are typically the first alleles to fail when it iscompromised resulting in allele drop-out. In the electropherogramsbelow, the template quality is excellent with peak balance running 85%to 100%, well above the acceptable range of 60% for accurate allelecalls.

1-29. (canceled)
 30. A method for collecting a biological sample thatcontains degradable biologically sourced analytes comprising: a)obtaining a device comprised of i) a sample collection member having ananalyte collection surface in the form of a pad of material mounted to astem; ii) a storage medium wherein the storage medium comprises at leastone stabilizing reagent that preserves at least one biological sampleanalyte for transport or storage; and iii) a storage medium holderhaving a means for removeably holding the storage medium in position anda means for pressing the pad onto the storage medium, for transfer of atleast a portion of the biological sample onto the storage medium; b)collecting a biological sample on the pad; c) operating the storagemedium holder to press the pad onto the storage medium; and d) releasingthe storage medium and pad.
 31. The method of claim 30, wherein themethod includes providing a visual delineation at an area of the storagemedium where transfer of the sample takes place.
 32. The method of claim30, wherein the means for pressing the pad onto the storage mediumprovides a reproducible pressure between the pad and the storage medium.33. A device for collecting a biological sample that contains degradablebiologically sourced analytes, the device comprising: i) a samplecollection member having an analyte collection surface in the form of apad of material mounted to a stem; ii) a storage medium wherein thestorage medium comprises at least one stabilizing reagent that preservesat least one biological sample analyte for transport or storage; andiii) a storage medium holder having a means for removeably holding thestorage medium in position and a means for pressing the pad onto thestorage medium, for transfer of at least a portion of the biologicalsample from the pad onto the storage medium.